In recent years, there have been developed numerous solid phase systems based on self-assembled monolayers (i.e., SAMs) from bi-functional molecules (i.e., “linkers”) that have gained ever more significance in analytical micro technology, wherein specific probe molecules are coupled, or, as the case may be, conjugated, onto the surface of a solid carrier, which can be confirmed with the help of suitable marking (for example, radio active, colored, or fluorescent marking).
For such systems, in analogy to electronic microchips, the term sensor chips has been adopted. In the case of the conjugation of biological molecules (so called “bioconjugation”) in such sensor chips, for example, with oligonucleotides or antibodies, one speaks of “Bio Chips.” Coupling on the surface of the carrier can be accomplished directly or indirectly. An example of indirect coupling is the coupling of a nucleic acid sequence to be tested by means of hybridisation onto an immobilized, complementary oligonucleotide used as a probe. In this case, the use of the probe has the additional advantage of the natural specificity of the interaction of biological macromolecules.
Typically, for the manufacture of sensor chips, surfaces of metal oxides, or, as the case may be, metalloid oxides, as, for example, aluminumoxide, quartz-glass, glass, are dipped in a solution of bi-functional molecules (so called “linkers”), which comprise, for example, a halosilane-(for example, chlorosilane-) or alkoxy silane group for coupling to the carrier surface so that a self-organized monolayer (SAM) forms. In this case, this SAM comprises a thickness of a few Ångstrom. The coupling of the linkers to the sample or probe molecules takes place by means of suitable further functional groups, for example, an amino or epoxy group. Suitable bi-functional linkers for coupling of a multitude of sample or probe molecules, particularly also those of biological origin, on a multitude of carrier surfaces, are known to one of ordinary skill in the art, see, for example, “Bioconjugate Techniques”, GT Hermanson, Academic Press, 1996.
A homogenious and chemically resistant layer can be obtained by this method only with much difficulty. Monofunctional silanes, which are suitable for solid carriers with metal oxide or metalloid oxide surfaces, for example, monochlorosilanes such as aminopropyl monochlorosilane, while they achieve real monolayers, can be split off from the surface merely by means of hot water. Trifunctional silanes, such as, for example, aminopropyltriethoxysilane, form cross-links in solution, which can lead to an inhomogeneous layer of silanes on the carrier surface. Homogeneous layering is not thereby achieved, which results in a loss in quality. In addition, there are also changes in character within a chip series.
A further disadvantage of this process in its practical application is the relatively long incubation time of silane solutions. For an efficient layering, silation times of about two hours are often necessary.
An object of the present invention is, departing from this state of the art, to fabricate a sensor chip with homogeneous layering based on a silane linker, as well as a process which makes such a layer quickly and reproducibly.